Aspartokinase (ATP:4-L-Aspartate-4-phosphotransferase [EC 2.7.2.4]) catalyses the conversion of aspartate and ATP to 4-phosphoaspartate and ADP. As shown in FIG. 1 for E. coli, aspartokinase is the first enzyme utilized in the biosynthetic pathway leading to lysine, threonine, and methionine. The biosynthesis of these nutritionally important amino acids is highly regulated. One mechanism for the regulation of this pathway is via the production of several isozymes of aspartokinase having different repressors and allosteric inhibitors. In both Escherichia coli and recently in Bacillus subtilis, three isozymes of aspartokinase differing in their sensitivity to repression and inhibition by lysine, threonine, methionine, and diaminopimelate have been identified. The three B. subtilis isozymes are feedback-inhibited by diaminopimelate, lysine, or threonine plus lysine, respectively (L. M. Graves, J. Bacteriol., 172, 218 (1990)). The lysine-sensitive aspartokinase II from B. subtilis has been purified to homogeneity by D. Moir et al., J. Biol. Chem., 252, 4648 (1977). The gene encoding this enzyme has also been cloned and sequenced, as reported by R. P. Bondaryk et al., J. Biol. Chem., 260, 592 (1985) and N. Y. Chen et al., J. Biol. Chem., 262, 8787 (1987).
Recently, F. J. Schendel et al. in J. Appl. Environ. Microbiol., 56, 963 (1990), identified homoserine auxotrophs and S-(2-aminoethyl)-cysteine (AEC) resistant mutants of a thermophilic methylotrophic Bacillus sp. which overproduce significant quantities of L-lysine at 50.degree. C. Such thermophilic methylotrophs may have advantages over other organisms for industrial use, as discussed by Al-Awadhi et al., Biotechnol. Bioeng., 36, 816, 821 (1990). In particular, the methylotrophic Bacillus MGA3 identified by F. J. Schendel et al., cited supra, may have significant advantages over other bacilli for the overproduction of lysine since it does not sporulate at high temperatures even under conditions of nutrient limitation, in contrast to lysine-producing mutants of B. licheniformis that sporulated when grown at temperatures greater than 40.degree. C. (H. Hagino et al., Biotechnol. Lett., 3, 425 (1981)).
Since both spore components, diaminopimelate and dipicolinic acid, are derived from the lysine biosynthetic pathway, as shown in FIG. 1, differences in the regulation of this pathway may occur between this thermophilic Bacillus sp. and other mesophilic bacilli. Therefore, a need exists to isolate and characterize the informational macromolecules (DNA and RNA) which function in the biosynthetic pathway to lysine, methionine and threonine in the thermotolerant Bacillus sp. MGA3. A further need exists to isolate and characterize the products, such as the enzymes, that function in these biosynthetic pathways. A further need exists to produce mutant varieties of said informational macromolecules, in order to improve the properties of the enzymes and other polypeptides encoded thereby, or to produce improved strains of thermotolerant, methylotrophic bacteria.